Advanced Medical Research Center, Yokohama City University, Yokohama 236-0004, Japan; R&D Headquarters, LION Corporation, Tokyo 132-0035, Japan.
Advanced Medical Research Center, Yokohama City University, Yokohama 236-0004, Japan.
J Proteomics. 2023 Sep 30;288:104976. doi: 10.1016/j.jprot.2023.104976. Epub 2023 Jul 22.
Although the microgravity (μ-g) environment that astronauts encounter during spaceflight can cause severe acute bone loss, the molecular mechanism of this bone loss remains unclear. To investigate the gravity-response proteins involved in bone metabolism, it is important to comprehensively determine which proteins exhibit differential abundance associated with mechanical stimuli. However, comprehensive proteomic analysis using small bone samples is difficult because protein extraction in mineralized bone tissue is inefficient. Here, we established a high-sensitivity analysis system for mouse bone proteins using data-independent acquisition mass spectrometry. This system successfully detected 40 proteins in the femoral diaphysis showing differential abundance between mice raised in a μ-g environment, where the bone mass was reduced by gravity unloading, and mice raised in an artificial 1-gravity environment on the International Space Station. Additionally, 22 proteins, including noncollagenous bone matrix proteins, showed similar abundance between the two groups in the mandible, where bone mass was unaltered due to mastication stimuli, suggesting that these proteins are responsive to mechanical stimuli. One of these proteins, SPARCL1, is suggested to promote osteoclastogenesis induced by receptor activator of nuclear factor-κB ligand. We expect these findings to lead to new insights into the mechanisms of bone metabolism induced by mechanical stimuli. SIGNIFICANCE: We aimed to investigate the gravity-response proteins involved in bone metabolism. To this end, we established a comprehensive analysis system for mouse bone proteins using data-independent acquisition mass spectrometry, which is particularly useful in comprehensively analyzing the bone proteome using small sample volumes. In addition, a comprehensive proteomic analysis of the femoral diaphysis and mandible, which exhibit different degrees of bone loss in mice raised on the International Space Station, identified proteins that respond to mechanical stimuli. SPARCL1, a mechanical stimulus-responsive protein, was consequently suggested to be involved in osteoclast differentiation associated with bone remodeling. Our findings represent an important step toward elucidating the molecular mechanism of bone metabolism induced by mechanical stimuli.
虽然宇航员在太空飞行中所经历的微重力（μ-g）环境会导致严重的急性骨丢失，但这种骨丢失的分子机制仍不清楚。为了研究与骨代谢相关的重力反应蛋白，全面确定哪些蛋白质与机械刺激相关的丰度差异很重要。然而，由于矿化骨组织中的蛋白质提取效率不高，使用小骨样本进行全面蛋白质组学分析是困难的。在这里，我们使用数据非依赖性采集质谱法建立了一种用于检测小鼠骨蛋白的高灵敏度分析系统。该系统成功地检测到在股骨骨干中 40 种蛋白的丰度存在差异，这些蛋白在微重力环境下培养的小鼠中，由于重力卸载导致骨量减少，而在国际空间站上的人工 1-g 环境中培养的小鼠中，这些蛋白的丰度也存在差异。此外，在咀嚼刺激导致骨量不变的下颌骨中，22 种蛋白（包括非胶原蛋白骨基质蛋白）在两组之间的丰度相似，这表明这些蛋白对机械刺激有反应。其中一种蛋白 SPARCL1 被认为可以促进核因子-κB 配体受体激活剂诱导的破骨细胞生成。我们希望这些发现能为机械刺激诱导的骨代谢机制提供新的见解。意义：我们旨在研究与骨代谢相关的重力反应蛋白。为此，我们使用数据非依赖性采集质谱法建立了一种用于检测小鼠骨蛋白的综合分析系统，该系统特别适用于使用小样本量全面分析骨蛋白质组。此外，对国际空间站上培养的小鼠股骨骨干和下颌骨进行全面蛋白质组学分析，这些骨骼在不同程度上发生了骨丢失，鉴定出了对机械刺激有反应的蛋白。机械刺激反应蛋白 SPARCL1 因此被认为参与了与骨重塑相关的破骨细胞分化。我们的发现为阐明机械刺激诱导的骨代谢分子机制迈出了重要一步。
